Cryobiology最新文献

{"title":"Morphological characteristics, mitochondrial oxidoreductase activity, and vascularization of human adipose tissues pre-and post-xenotransplantation into BALB/c female nude mice","authors":"Banafsheh Heidari ,&nbsp;Negar Eidi ,&nbsp;Pejman Mortazavi ,&nbsp;Zahra Saffarian ,&nbsp;Azadeh Soltani ,&nbsp;Parvin Mansouri ,&nbsp;Seyed Mehdi Tabaie","doi":"10.1016/j.cryobiol.2025.105248","DOIUrl":"10.1016/j.cryobiol.2025.105248","url":null,"abstract":"<div><div>This study aimed to evaluate the effect of different cryoprotectants on the histomorphological characteristics, oil ratio (OR) indices, mitochondrial oxidoreductase activities, and vascularization of human adipose tissues after thawing (pre-xenotransplantation), one month and three months post-xenotransplantation. For this purpose, we froze human adipose tissues in five experimental groups, including NCG (PBS without any cryoprotectants), FG1 (4 % DMSO+6 % Trehalose), FG2 (6 % Trehalose+2 % DMSO+9 % FBS), FG3 (2 % DMSO+4 % Trehalose+3 % Sucrose+9 % FBS), and FG4 (4 % DMSO+3 % Trehalose+3 % Sucrose) and compared them with the positive control group (PCG or fresh adipose tissue). Adipose tissues were frozen with a cooling rate of −1 °C/min and stored in a freezer at −20 °C until use. Three months after freezing, human adipose tissues were xenotransplanted into the BALB/c nude mice for one and three months. Then, xenografts were removed and evaluated for histopathological properties, OR indices, mitochondrial activities, and vascularization. After thawing (pre-xenotransplantation) and post-xenotransplantation (one- and three-month after xenografting), the NCG group demonstrated the most significant tissue damage including necrosis, multiple cysts, inflammatory cell invasion, cell rupture, fibrosis, necrosis and ECM disintegration (<em>p ≤</em> 0.05). Pre-xenotransplantation, one- and three-month after grafting, the best structural integrity of adipose tissues with the least degeneration of adipocytes was detected in the FG2 and FG3 groups (<em>p ≤</em> 0.05). Pre- and post-xenotransplantation, the highest and least OR indices and mitochondrial oxidoreductase activities were demonstrated in the NCG and FG2 groups, respectively (<em>p ≤</em> 0.05). The least VEGF and CD31 expressions were shown in the NCG group at pre- and post-transplantation (<em>p ≤</em> 0.05). The highest CD31 and VEGF expressions were detected in the FG3 and FG2 groups after thawing and grafting (both one- and three-month) (<em>p ≤</em> 0.05).</div></div>","PeriodicalId":10897,"journal":{"name":"Cryobiology","volume":"119 ","pages":"Article 105248"},"PeriodicalIF":2.3,"publicationDate":"2025-05-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143911627","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Optimization of freezing and thawing protocols for human ovarian tissue cryopreservation through thermophysical characterisation of freezing medium by differential scanning calorimetry","authors":"Fiot Mélanie ,&nbsp;Martinot Emmanuelle ,&nbsp;Travens Lincoln ,&nbsp;Dollet Sandra ,&nbsp;Radosevic-Robin Nina ,&nbsp;Lucas Cécily ,&nbsp;Chaput Laure ,&nbsp;Pereira Bruno ,&nbsp;Gremeau Anne-Sophie ,&nbsp;Hochet Emmanuel ,&nbsp;Gavin-Plagne Lucie ,&nbsp;Baudot Anne ,&nbsp;Brugnon Florence ,&nbsp;Marteil Gaëlle","doi":"10.1016/j.cryobiol.2025.105245","DOIUrl":"10.1016/j.cryobiol.2025.105245","url":null,"abstract":"<div><div>Fertility preservation should be offered to patients facing gonadotoxic therapy. The method for preserving prepubescent girls' fertility, which is also suitable for women, is ovarian tissue cryopreservation (OTC). Although 200 births have been reported worldwide with this approach, significant improvements are needed. The literature indeed reports numerous protocols for freezing and thawing ovarian tissue, with no clear rationale for selection criteria. This study aims to optimize human OTC protocols by characterizing the thermodynamic properties of freezing medium. The freezing medium associated with most live births after autograft (Leibovitz L-15 medium with 4 mg/mL human serum albumin (HSA), 1.5M DMSO, and 0.1M sucrose) was characterized using differential scanning calorimetry. We obtained −120.49 °C for glass transition temperature (Tg’), −20 °C for crystallization temperature when cooling at 2.5 °C/min (Tc) and −4.11 °C for melting temperature (Tm). With these parameters, we optimized a freezing protocol in a programmable freezer (Nano-Digitcool, Cryo Bio System) and a thawing protocol. The freezing curve was as follows: 5 min at 4 °C, 1 °C/min to −7 °C, seeding: 60 °C/min to −32 °C, and 10 °C/min to −15 °C, 0.3 °C/min to −40 °C, 10 °C/min to −140 °C. The thawing protocol consisted in a 3.5-min step in a cold chamber to reach slowly Tg', limiting thermal and mechanical shocks, and then a 2-min incubation at 37 °C to quickly reach Tm. Ovarian tissue frozen-thawed according to these protocols had a similar quality to that of fresh tissue and could resume folliculogenesis during organotypic culture. Our study will contribute to improve human OTC and optimize women fertility preservation.</div></div>","PeriodicalId":10897,"journal":{"name":"Cryobiology","volume":"119 ","pages":"Article 105245"},"PeriodicalIF":2.3,"publicationDate":"2025-05-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143895597","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Influence of extenders and antioxidants on Nile tilapia (Oreochromis niloticus) sperm parameters","authors":"Vinícius Wagner Silva ,&nbsp;Myrian Megumy Tsunokawa Hidalgo ,&nbsp;Renan José Casarotto Appel ,&nbsp;Karine Nicole Siqueira ,&nbsp;José Beirão ,&nbsp;Laurival Antônio Vilas-Boas ,&nbsp;Lucienne Garcia Pretto-Giordano ,&nbsp;Rajesh Joshi ,&nbsp;Jorge Manuel de Oliveira Fernandes ,&nbsp;Maria Isabel Mello Martins","doi":"10.1016/j.cryobiol.2025.105253","DOIUrl":"10.1016/j.cryobiol.2025.105253","url":null,"abstract":"<div><div>This study aimed to evaluate the effects of different dilutions of extenders (ionic, glucose, and bicine) and antioxidant concentrations on Nile tilapia sperm kinetics and membrane integrity parameters (MI). Antioxidants tested included taurine (1, 5, and 50 mmol), uric acid (0.5, 1, and 5 mmol), reduced glutathione (1, 5, and 10 mmol), and ascorbic acid (1, 10, and 50 mmol). Initially, ejaculates from five Nile tilapia were collected, evaluated, and diluted in a ratio of 1:3 into three aliquots, each with ionic, glucose, or bicine extenders. The samples were stored at 4 °C for 60 min and evaluated for kinetics and MI. The ionic extender showed higher MI and progressive motility. Then, ejaculates of four fish were collected to test each antioxidant. The samples were diluted in a ratio of 1:3, into a control group (without antioxidants) and the three concentrations in an ionic extender plus methanol 10 %. The samples were frozen/thawed and evaluated for kinetics and MI. Taurine at 1 mmol showed higher total motility, cells with medium velocity, and lower static cells. No differences were found in uric acid concentrations tested. Reduced glutathione treatments resulted in higher membrane damage, number of static cells, and lower total motility. Ascorbic acid addition decreased total motility and increased the number of static cells. Therefore, the addition of 1 mmol taurine was suitable for Nile tilapia sperm cryopreservation. Uric acid in tested concentrations maintained the sperm parameters, while reduced glutathione and ascorbic acid decreased sperm quality.</div></div>","PeriodicalId":10897,"journal":{"name":"Cryobiology","volume":"119 ","pages":"Article 105253"},"PeriodicalIF":2.3,"publicationDate":"2025-05-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143895561","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"β-Carotene addition into a Tris-based diluent improves Hu ram sperm parameters after cryopreservation","authors":"Liuming Zhang ,&nbsp;Caiyu Jiang ,&nbsp;Yuxuan Sun ,&nbsp;Fuhao Chen ,&nbsp;Jian Wang ,&nbsp;Yongjun Li","doi":"10.1016/j.cryobiol.2025.105254","DOIUrl":"10.1016/j.cryobiol.2025.105254","url":null,"abstract":"<div><div>The effect of β-Carotene on ram semen cryopreservation has not been previously evaluated. The present research aimed to investigate the impact of different concentrations of β-Carotene in the diluent on the quality of ram semen and to analyze its antioxidant effects by the addition of tert-butyl hydroperoxide (TBHP) during cryopreservation. Semen samples were diluted with a Tris-based extender containing the β-Carotene (0, 10, 20, 30 and 40 mg/L). The motility and biokinetic characteristics, membrane and acrosome integrity, related parameters of oxidative stress, mitochondrial membrane potential (MMP) and apoptosis rate were measured after the cryo-preservation. The results indicated that the 20 mg/L β-Carotene group significantly improved total motility (TM), progressive motility (PM), average motion degree (MAD), membrane and acrosome integrity, total antioxidant capacity (T-AOC), superoxide dismutase (SOD) and catalase (CAT) compared to the other groups (P &lt; 0.05). Additionally, this group significantly reduced reactive oxygen species (ROS) and malondialdehyde (MDA) content compared to the control group (P &lt; 0.05). Furthermore, the TM, PM, wobble (WOB), MAD, membrane and acrosome integrity, CAT, SOD, T-AOC and MMP in the 100 μM TBHP+20 mg/ml β-Carotene group were higher than those in the 100 μM TBHP group (P &lt; 0.05). The combined supplementation group also significantly decreased the ROS, MDA, the protein level of Cytochrome C and sperm apoptosis rates (P &lt; 0.05). Therefore, 20 mg/L is identified as the optimal concentration for the cryopreservation of ram semen, and β-Carotene can improve the oxidative stress damage during the frozen-thawed process by enhancing the antioxidant capacity of sperm and maintaining mitochondrial function.</div></div>","PeriodicalId":10897,"journal":{"name":"Cryobiology","volume":"119 ","pages":"Article 105254"},"PeriodicalIF":2.3,"publicationDate":"2025-05-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143895473","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Low-temperature stress response: A transcriptomic study of the WRKY family in Prunus davidiana","authors":"Meiling Guo ,&nbsp;Rongjun Pan ,&nbsp;Zhenjing Chu ,&nbsp;Wenxian Gai ,&nbsp;Xuqiang Qiao ,&nbsp;Meixia Liang","doi":"10.1016/j.cryobiol.2025.105252","DOIUrl":"10.1016/j.cryobiol.2025.105252","url":null,"abstract":"<div><div>Low temperature is a crucial environmental factor affecting peach tree growth. The WRKY transcription factor family plays a vital role in plants' responses to low-temperature stress. Understanding the expression patterns and functions of WRKY genes in peach trees under low-temperature stress is essential for uncovering the molecular mechanisms of cold tolerance and guiding the breeding of new cold-tolerant peach varieties. This study analyzed the transcriptome of cold-tolerant peach varieties following low-temperature treatment. Bioinformatics and quantitative real-time PCR were used to systematically analyze the structure, function, and expression characteristics of WRKY family members associated with low-temperature tolerance in peach trees. Results revealed complex transcriptional responses in peach trees under low-temperature stress, with 7029 differentially expressed genes identified and significantly enriched in multiple Gene Ontology terms and Kyoto Encyclopedia of Genes and Genomes pathways, indicating the diverse mechanisms of low-temperature response. Among the transcription factor families, the ERF, MYB, bHLH, NAC, and WRKY families showed significant responses to low-temperature stress. <em>PpWRKY58</em> exhibited robust and specific upregulation under low-temperature stress, and yeast transformation showed enhanced activity under low-temperature conditions. This suggests a potential role for <em>PpWRKY58</em> in peach cold hardiness. While WRKY family involvement in plant responses to low-temperature stress is known, our study reveals the specific function of <em>PpWRKY58</em> in the peach low-temperature response, offering new insights into peach cold tolerance mechanisms, and suggesting <em>PpWRKY58</em> as a potential target for improving peach cold hardiness.</div></div>","PeriodicalId":10897,"journal":{"name":"Cryobiology","volume":"119 ","pages":"Article 105252"},"PeriodicalIF":2.3,"publicationDate":"2025-04-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143890703","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Chromosomal integrity of freeze-dried karyoplasts in mice","authors":"Hirokazu Kusakabe","doi":"10.1016/j.cryobiol.2025.105247","DOIUrl":"10.1016/j.cryobiol.2025.105247","url":null,"abstract":"<div><div>In mammalian species, there is currently no way to preserve mature oocytes at supra-zero temperatures without cryostorage. Metaphase II (MII) oocytes freeze-dried in any medium or solution cannot be revived after rehydration. Therefore, the injurious effects to chromosomes of freeze-drying MII oocytes have not been reported. The aim of this study was to examine the chromosomal integrity of freeze-dried MII oocytes in mice. Spindle apparatuses with small amounts of cytoplasm, “karyoplasts” so-called, were removed from MII mouse oocytes. Before freeze-drying (FD), the karyoplasts were incubated at 4 °C for up to 2 days (pre-FD incubation) in EGTA/Tris-HCl buffered solution supplemented with 20 μmol/l γ-tocotrienol. After freeze-drying, the freeze-dried karyoplasts were rehydrated and microinjected into enucleated MII oocytes. Parthenogenetic activation of the reconstructed oocytes was performed to analyze the chromosomes at the first cleavage metaphase. A portion of normally activated oocytes that had been injected with fresh karyoplasts (51 %) and karyoplasts freeze-dried after pre-FD incubation for 8 h to 2 d (27 %–29 %) exhibited normal chromosome constitution. Insufficient pre-FD incubation (0–4 h) caused severe chromosomal damage. By contrast, almost all parthenogenetic embryos (98 %) reconstructed via fusion of fresh karyoplasts and enucleated oocytes maintained normal chromosome constitution. Some MII oocytes reconstructed and activated parthenogenetically using freeze-dried karyoplasts could develop the first cleavage metaphase (67–80 %) while retaining chromosome stability (3–29 %). Further improvements in FD procedures should enhance the chromosomal integrity of freeze-dried karyoplasts.</div></div>","PeriodicalId":10897,"journal":{"name":"Cryobiology","volume":"119 ","pages":"Article 105247"},"PeriodicalIF":2.3,"publicationDate":"2025-04-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143890704","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"The effects of various synthetic polymers on oxidative stress in ovarian tissue subjected to cryoprotectant exposure and vitrification","authors":"Ebrahim Asadi ,&nbsp;Atefeh Najafi ,&nbsp;James D. Benson","doi":"10.1016/j.cryobiol.2025.105243","DOIUrl":"10.1016/j.cryobiol.2025.105243","url":null,"abstract":"<div><div>To enhance the post-thaw viability of ovarian tissue, cryopreservation methods are continually being refined. Therefore, we aimed to explore whether ovarian tissue vitrification in closed high-security tubes could be improved with ice-blocking polymers. Bovine ovarian fragments (n = 10) were exposed to varying concentrations of a vitrification solution containing glycerol and ethylene glycol (VS1 (54 % (w/v)), VS2 (57 % (w/v)), VS3 (60 % (w/v))), supplemented with synthetic polymers (Super cool X-1000, Super cool Z-1000, and polyvinylpyrrolidone (PVP K-12)), with (VVS1, VVS2, VVS3) or without subsequent vitrification. In the second phase (n = 6), we explored the individual effects of each polymer. A lower percentage of abnormal follicles was observed in the fresh and VVS2 groups compared to other groups. Additionally, the fresh, VVS2, and VVS3 groups exhibited less tissue fibrosis than the VVS1 group. Higher tissue viability, total antioxidant levels, and lower reactive oxygen species (ROS) levels were observed in the fresh and VVS2 groups. In groups exposed to CPA, there was an increase in ROS and a decrease in antioxidants compared to the fresh group. We also observed higher tissue viability and improved follicular morphology in the vitrification solution supplemented with Super cool X-1000, either alone or in combination with PVP K-12 and Super cool Z-1000. Lower ROS were found in the group supplemented with PVP K-12, either alone or in combination with Super cool X-1000 and Super cool Z-1000. In conclusion, combining a closed vitrification system with an optimized concentration of cryoprotective agents and synthetic polymers improved bovine ovarian tissue vitrification outcomes including post thaw viability and follicular morphology when compared to standard cryopreservation protocols.</div></div>","PeriodicalId":10897,"journal":{"name":"Cryobiology","volume":"119 ","pages":"Article 105243"},"PeriodicalIF":2.3,"publicationDate":"2025-04-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143859245","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Conjugates of gold nanoparticles and antifreeze protein III for cryopreservation of cells and tissues","authors":"Mariia Yukhta ,&nbsp;Iryna Bespalova ,&nbsp;Oleksandra Hubenia ,&nbsp;Ido Braslavsky ,&nbsp;Boris N. Chichkov ,&nbsp;Oleksandr Gryshkov","doi":"10.1016/j.cryobiol.2025.105246","DOIUrl":"10.1016/j.cryobiol.2025.105246","url":null,"abstract":"<div><div>The development of non-toxic cryoprotectants is crucial for advancing fields such as regenerative medicine, cell therapy and tissue engineering, where the preservation of the viability of cells, tissues and organs during cryopreservation is essential. This interdisciplinary effort involves areas such as cryobiology, nanotechnology, biochemistry and material science to create more efficient and safer cryoprotective solutions. This study explores the development and application of gold nanoparticles (AuNPs) conjugated with antifreeze protein III (AFPIII) for improving the cryopreservation of bone marrow stem cells (bMSCs) encapsulated in alginate macrospheres (AMSs). Two types of AuNPs, stabilized with citrate (CitAuNPs) and BSPP (BSPPAuNPs), were functionalized with AFPIII using both covalent and non-covalent conjugation methods and were characterized for their size, surface charge and protein layer thickness. The cytotoxicity assays indicated that both types of AuNPs and their AFPIII conjugates had no adverse effects on bMSC viability and proliferation over 48 h, demonstrating their non-toxicity. Furthermore, the cryopreservation of bMSC-contained AMSs revealed that the covalent BSPPAuNPs-AFPIII conjugate provided superior preservation of cell viability and metabolic activity, outperforming both non-covalent conjugates and individual components. Cryomicroscopic analysis revealed that AFPIII altered ice crystal formation, promoting smaller, multidirectional crystals, which minimized cellular damage during freezing. The covalent BSPPAuNPs-AFPIII conjugate exhibited superior cryoprotective effects, preserving cell viability and function better than the non-covalent CitAuNPs-AFPIII conjugate. These findings suggest that AuNPs-AFPIII conjugates, particularly the covalent BSPPAuNPs-AFPIII complex, hold great promise for improving cell and tissue cryopreservation protocols.</div></div>","PeriodicalId":10897,"journal":{"name":"Cryobiology","volume":"119 ","pages":"Article 105246"},"PeriodicalIF":2.3,"publicationDate":"2025-04-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143848074","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Cryopreservation of spermatogonia of the giant freshwater prawn, Macrobrachium rosenbergii, using slow and ultra-rapid freezing","authors":"Tomoyuki Okutsu ,&nbsp;Natthida Rakbanjong ,&nbsp;Shinya Shikina ,&nbsp;Misako Miwa ,&nbsp;Monwadee Wonglapsuwan","doi":"10.1016/j.cryobiol.2025.105242","DOIUrl":"10.1016/j.cryobiol.2025.105242","url":null,"abstract":"<div><div>The giant freshwater prawn, <em>Macrobrachium rosenbergii</em>, is an important species that is widely raised in tropical and subtropical regions. To develop techniques for preserving valuable traits in this species, we have devised protocols for cryopreserving spermatogonia—progenitor cells that give rise to spermatozoa through spermatogenesis—using slow freezing and ultra-rapid freezing techniques. Optimization of cryoprotectants and their concentrations revealed that 10 % dimethyl sulfoxide (ME₂SO) is effective as a cryoprotectant in both methods. The optimal time for equilibration to the cryoprotectant before slow freezing was determined to be 15 min. Thawing temperature had a significant effect on cell recovery and viability, both being markedly higher when cells were thawed at 10 °C than at 27 °C. After long-term storage in liquid nitrogen, cells preserved by ultra-rapid freezing exhibited higher recovery and viability rates than those preserved using the slow freezing method, with recovery rates of up to 86.4 % ± 5.9 % at 6 months. Therefore, using 10 % ME₂SO with the ultra-rapid freezing method is recommended as the optimal protocol for long-term cryopreservation of <em>M. rosenbergii</em> spermatogonia.</div></div>","PeriodicalId":10897,"journal":{"name":"Cryobiology","volume":"119 ","pages":"Article 105242"},"PeriodicalIF":2.3,"publicationDate":"2025-04-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143828421","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Dynamics of amino acids in the shoot tips of Pogostemon yatabeanus during cryopreservation by droplet-vitrification method and its modifications","authors":"Byeongchan Choi ,&nbsp;Hyoeun Lee ,&nbsp;Elena Popova ,&nbsp;Man-Jeong Paik ,&nbsp;Haenghoon Kim","doi":"10.1016/j.cryobiol.2025.105244","DOIUrl":"10.1016/j.cryobiol.2025.105244","url":null,"abstract":"<div><div>Cryopreservation offers effective long-term conservation of the endangered plant species with non-orthodox or unavailable seeds. Shoot tips of <em>Pogostemon yatabeanus</em>, an endemic Korean species, were successfully cryopreserved via the optimized droplet-vitrification (DV) method which included 10 % sucrose preculture, osmoprotection with 17.5 % glycerol + 17.5 % sucrose, vitrification solution (33.3 % glycerol + 13.3 % dimethyl sulfoxide + 13.3 % ethylene glycol + 20.1 % sucrose) treatment and cooling-rewarming using aluminum foil strips. Regrowth was performed in three steps, starting with the ammonium-free medium with growth regulators, followed by full-strength medium with and without growth regulators in the second and the third steps. The contents of 17 amino acids (AA) in shoot tips were analyzed using gas-chromatography-mass-spectrometry (GC-MS/MS) at each step of the optimum DV protocol and in the following modifications: excluding or modifying preculture or osmoprotection, varying vitrification solution composition, cooling-rewarming in vials, change of regrowth conditions. In the optimum DV procedure, the total content of AA increased by 20–40 % at preculture and osmoprotection, dropped to 6 % of their level in freshly excised shoot tips after the first five days of regrowth, and raised again after two weeks of recovery. The contents of alanine, glycine, β-alanine, serine, and γ-aminobutyric acid were maximized during osmoprotection and vitrification solution treatments (6.27–29.75-fold compared to untreated control). In particular, alanine showed a 30-fold peak during osmoprotection. Excluding preculture or osmoprotection decreased total and individual AA concentrations by 40–70 %. Meanwhile, a high (25 %) sucrose concentration during preculture increased AA' contents almost twice compared to the optimum protocol. Modifications of vitrification solutions caused 20–30 % variations in AA contents. Therefore, AA metabolism is essential in shoot tips’ response to osmotic and freezing stresses and regeneration</div></div>","PeriodicalId":10897,"journal":{"name":"Cryobiology","volume":"119 ","pages":"Article 105244"},"PeriodicalIF":2.3,"publicationDate":"2025-04-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143826379","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}